Abstract
comC is a Bacillus subtilis gene required for the development of genetic competence. We have cloned a fragment from the B. subtilis chromosome that carries comC and contains all the information required to complement a Tn917lac insertion in comC. Genetic tests further localized comC to a 2.0-kilobase HindIII fragment. Northern (RNA) blotting experiments revealed that an 800-base-pair comC-specific transcript appeared at the time of transition from exponential to stationary phase during growth through the competence regimen. The DNA sequence of the comC region revealed two open reading frames (ORFs), transcribed in the same direction. The upstream ORF encoded a protein with apparent sequence similarity to the folC gene of Escherichia coli. Insertion of a chloramphenicol resistance determinant into this ORF and integration of the disrupted construct into the bacterial chromosome by replacement did not result in competence deficiency. The downstream ORF, which contained the Tn917lac insertion that resulted in a lack of competence, is therefore the comC gene. The predicted protein product of comC consisted of 248 amino acid residues and was quite hydrophobic. The comC gene product was not required for the expression of any other com genes tested, and this fact, together with the marked hydrophobicity of ComC, suggests that it may be a component of the DNA-processing apparatus of competent cells.
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- Albano M., Breitling R., Dubnau D. A. Nucleotide sequence and genetic organization of the Bacillus subtilis comG operon. J Bacteriol. 1989 Oct;171(10):5386–5404. doi: 10.1128/jb.171.10.5386-5404.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Albano M., Dubnau D. A. Cloning and characterization of a cluster of linked Bacillus subtilis late competence mutations. J Bacteriol. 1989 Oct;171(10):5376–5385. doi: 10.1128/jb.171.10.5376-5385.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Albano M., Hahn J., Dubnau D. Expression of competence genes in Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3110–3117. doi: 10.1128/jb.169.7.3110-3117.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Anagnostopoulos C., Spizizen J. REQUIREMENTS FOR TRANSFORMATION IN BACILLUS SUBTILIS. J Bacteriol. 1961 May;81(5):741–746. doi: 10.1128/jb.81.5.741-746.1961. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bognar A. L., Osborne C., Shane B. Primary structure of the Escherichia coli folC gene and its folylpolyglutamate synthetase-dihydrofolate synthetase product and regulation of expression by an upstream gene. J Biol Chem. 1987 Sep 5;262(25):12337–12343. [PubMed] [Google Scholar]
- Bognar A. L., Osborne C., Shane B., Singer S. C., Ferone R. Folylpoly-gamma-glutamate synthetase-dihydrofolate synthetase. Cloning and high expression of the Escherichia coli folC gene and purification and properties of the gene product. J Biol Chem. 1985 May 10;260(9):5625–5630. [PubMed] [Google Scholar]
- Cahn F. H., Fox M. S. Fractionation of transformable bacteria from ocompetent cultures of Bacillus subtilis on renografin gradients. J Bacteriol. 1968 Mar;95(3):867–875. doi: 10.1128/jb.95.3.867-875.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dale R. M., McClure B. A., Houchins J. P. A rapid single-stranded cloning strategy for producing a sequential series of overlapping clones for use in DNA sequencing: application to sequencing the corn mitochondrial 18 S rDNA. Plasmid. 1985 Jan;13(1):31–40. doi: 10.1016/0147-619x(85)90053-8. [DOI] [PubMed] [Google Scholar]
- Dubnau D., Davidoff-Abelson R. Fate of transforming DNA following uptake by competent Bacillus subtilis. I. Formation and properties of the donor-recipient complex. J Mol Biol. 1971 Mar 14;56(2):209–221. doi: 10.1016/0022-2836(71)90460-8. [DOI] [PubMed] [Google Scholar]
- Dubnau D., Davidoff-Abelson R., Smith I. Transformation and transduction in Bacillus subtilis: evidence for separate modes of recombinant formation. J Mol Biol. 1969 Oct 28;45(2):155–179. doi: 10.1016/0022-2836(69)90097-7. [DOI] [PubMed] [Google Scholar]
- Engelman D. M., Steitz T. A., Goldman A. Identifying nonpolar transbilayer helices in amino acid sequences of membrane proteins. Annu Rev Biophys Biophys Chem. 1986;15:321–353. doi: 10.1146/annurev.bb.15.060186.001541. [DOI] [PubMed] [Google Scholar]
- Gaur N. K., Dubnau E., Smith I. Characterization of a cloned Bacillus subtilis gene that inhibits sporulation in multiple copies. J Bacteriol. 1986 Nov;168(2):860–869. doi: 10.1128/jb.168.2.860-869.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gryczan T. J., Hahn J., Contente S., Dubnau D. Replication and incompatibility properties of plasmid pE194 in Bacillus subtilis. J Bacteriol. 1982 Nov;152(2):722–735. doi: 10.1128/jb.152.2.722-735.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guillen N., Weinrauch Y., Dubnau D. A. Cloning and characterization of the regulatory Bacillus subtilis competence genes comA and comB. J Bacteriol. 1989 Oct;171(10):5354–5361. doi: 10.1128/jb.171.10.5354-5361.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hadden C., Nester E. W. Purification of competent cells in the Bacillus subtilis transformation system. J Bacteriol. 1968 Mar;95(3):876–885. doi: 10.1128/jb.95.3.876-885.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hahn J., Albano M., Dubnau D. Isolation and characterization of Tn917lac-generated competence mutants of Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3104–3109. doi: 10.1128/jb.169.7.3104-3109.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Israeli-Reches M., Weinrauch Y., Dubnau D. Evolutionary relationships of the Bacillus licheniformis macrolide-lincosamide-streptogramin B resistance elements. Mol Gen Genet. 1984;194(3):362–367. doi: 10.1007/BF00425545. [DOI] [PubMed] [Google Scholar]
- Lipman D. J., Pearson W. R. Rapid and sensitive protein similarity searches. Science. 1985 Mar 22;227(4693):1435–1441. doi: 10.1126/science.2983426. [DOI] [PubMed] [Google Scholar]
- Nonet M. L., Marvel C. C., Tolan D. R. The hisT-purF region of the Escherichia coli K-12 chromosome. Identification of additional genes of the hisT and purF operons. J Biol Chem. 1987 Sep 5;262(25):12209–12217. [PubMed] [Google Scholar]
- Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
- Shaw J. H., Clewell D. B. Complete nucleotide sequence of macrolide-lincosamide-streptogramin B-resistance transposon Tn917 in Streptococcus faecalis. J Bacteriol. 1985 Nov;164(2):782–796. doi: 10.1128/jb.164.2.782-796.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shine J., Dalgarno L. The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1342–1346. doi: 10.1073/pnas.71.4.1342. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
- Ulmanen I., Lundström K., Lehtovaara P., Sarvas M., Ruohonen M., Palva I. Transcription and translation of foreign genes in Bacillus subtilis by the aid of a secretion vector. J Bacteriol. 1985 Apr;162(1):176–182. doi: 10.1128/jb.162.1.176-182.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weinrauch Y., Guillen N., Dubnau D. A. Sequence and transcription mapping of Bacillus subtilis competence genes comB and comA, one of which is related to a family of bacterial regulatory determinants. J Bacteriol. 1989 Oct;171(10):5362–5375. doi: 10.1128/jb.171.10.5362-5375.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Youngman P., Perkins J. B., Losick R. A novel method for the rapid cloning in Escherichia coli of Bacillus subtilis chromosomal DNA adjacent to Tn917 insertions. Mol Gen Genet. 1984;195(3):424–433. doi: 10.1007/BF00341443. [DOI] [PubMed] [Google Scholar]